This invention relates to a method of and an apparatus for separating lymphocytes from a lymphocyte-containing suspension by filtration.
By the term "lymphocyte-containing suspension" used herein is meant blood, ascites, bone marrow and other lymphocyte- or lymphocyte precursor cell-containing body fluids. This term should also be interpreted as including physically, chemically and/or biologically treated blood and other body fluids such as, for example, blood diluted with a physiological solution, erythrocyte agglutinant-(such as dextran or hydroxyethyl-starch)-incorporated blood, a buffy coat and other lymphocyte-containing suspension layers prepared by centifugation or cell-electrophoresis.
In recent years lymphocytes collected from blood have been in frequent use in the field of immunology. For example, lymphocytes are used for immunological studies directed to histocompatibility antigens and cellular immunity. Lymphocytes are also used for determining lymphocyte blastotransformation or for determining the proportion of T-cells to B cells, and for trying to separate helper T-cells and suppressor T cells from T cells. Furthermore, lymphocyte component transfusions have been studied in great detail.
Various processes have been employed for carrying out lymphocyte separation. Typical separation processes include, for example, a density gradient centrifugation process such as the Ficoll-Conray process, an anticoagulant incorporation-sedimentation-(or centrifugation)-adhering process and a cell electrophoresis process.
The centrifugation process involves, for example, placing a plurality of liquids of different densities, one upon another, in a vessel to form density gradient superposed layers; placing blood on the top liquid layer; and then centrifuging the superposed layers thereby to separate the blood into several layers.
The anticoagulant incorporation-sedimentation-(or centrifugation)-adhering process involves incorporating an anticoagulant into a lymphocyte-containing suspension, subjecting the anticoagulant-incorporated suspension to sedimentation or centrifugation to separate the suspension into an erythrocyte sediment and a leukocyte-containing liquid layer, then introducing the leukocyte-containing liquid layer into a column having packed therein a mass of fibers such as glass wool, cotton or nylon wool, wherein the liquid is maintained at a temperature of 37.degree. C. for a period of approximately 30 minutes to entrap granulocytes and monocytes in the mass of fibers, and finally collecting lymphocytes.
The above-mentioned conventional processes have several disadvantages as will now be described. The density gradient centrifugation process requires a substantial period of time and skill to carry out the centrifugation step completely. When this process is employed, lymphocytes are likely to be undesirably changed during such a long period of time and, furthermore, to be damaged by the centrifugation and repeated washing operations. The density gradient centrifugation process further requires the use of a plurality of expensive apparatuses. When the lymphocytes collected by this centrifugation process are used for transfusion, additives such as a Ficoll-Conray solution or gum arabic are required to be removed from the collected lymphocytes.
In the anticoagulant incorporation-sedimentation-adhering process, a substantial period of time is required to complete separation of lymphocytes from the lymphocyte-containing suspension and it is also difficult to collect a lymphocyte-rich suspension with a high yield. The cell electrophoresis process requires the use of an expensive apparatus and is therefore disadvantageous.